Mechanisms of cell death are usually classified into two pathways, apoptosis and necrosis. However, it has been proposed by the American Society of Toxicologic Pathologists that the term oncosis, with its root meaning of "swelling" be used as the alternate descriptor of cell death occurring by non-apoptotic pathways. Necrosis more accurately describes the consequences of oncotic cell death, usually the death of a large number of cells which results in moderate to severe tissue injury. Oncosis is a form of cell death that typically occurs in response to toxic injury, including that induced by chemical exposure and reactive oxygen species (ROS). ROS are involved in the initiation and progression of a variety of human diseases and toxicities associated with chemical exposure. An understanding of the factors that regulate the cellular response to ROS and of the molecular mechanisms by which they interact with cellular constituents, and the consequences of such interactions, are important fundamental goals of biomedical research. The generation of ROS has been implicated in the pathogenesis of many pathological conditions. We have been using an in vitro model, in which H202 is generated in situ following the addition of TGHQ to LLC-PK1 cells, to investigate the cellular and molecular response of renal proximal tubule epithelial cells to oxidant-induced injury. Rigid controls function to prevent repeated rounds of DNA replication (S-phase arrest) without intervening mitoses, or the initiation of mitosis (G2M arrest) before DNA replication is complete ("mitotic catastrophe"). Loss of these cell cycle checkpoints after DNA damage may permit premature entry into mitosis. Our Preliminary data indicate that ROS-induced ERK activation contributes to oncotic cell death of LLC-PK1 cells by a mechanism that involves premature chromatin condensatior (PCC) and premature entry into mitosis. Four Specific Aims are proposed to test four inter-related hypotheses. (1) ERK activation is coupled to PCC and mitotic catastrophe via the activation of downstream histone H3 kinases. (2) PARP mediated ADP-ribosylation of histones facilitates histone H3 phosphorylation, and these post-translational modifications, perhaps in combination with additional modifications, are required for PCC and mitotic catastrophe. (3) ROS interfere with one or more components of the DNA damage check point system, driving the cells into premature mitosis, and subsequently death by mitotic catastrophe. (4) ROS induce the inappropriate nuclear translocation of cell cycle regulators, promoting premature mitosis. Our data, and that of others, indicate that responses to stress that usually result in oncotic cell death (and tissue necrosis) can indeed be manipulated, at the genetic and pharmacological level, to produce a potentially favorable (survivable) tissue response. Experiments proposed in the present application are designed to address this possibility. Basic knowledge of the mechanisms by which ROS induce cell death may yield strategies for clinical interventions in pathologies in which ROS play a prominent role